The ultimate goal of this proposal is to determine the cause(s) of impaired glucose tolerance leading to type 2 diabetes. An improved understanding of the underlying pathophysiology of the early events should provide a rational basis for the development of improved methods of early diagnosis and ultimately treatments for type 2 diabetes. Increased availability of lipid substrates, particularly plasma non-esterified fatty acids and intracellular triglyceride stores have been linked to many aspects of the insulin resistance syndrome including obesity, dyslipidemia and Type 2 Diabetes. The pancreatic beta cell response to nutrient oversupply and obesity associated insulin resistance is compensatory insulin hypersecretion in order to maintain euglycemia. Diabetes, and early states of glucose intolerance only develop in those who develop beta cell dysfunction. Although beta cell mass has been found to decline by ~50% in those with prediabetic states of glucose intolerance, controversy exists as to whether defects in the beta cell secretory response is due to deficient synthetic machinery or functional defects in glucose sensing and insulin secretion. This proposal will test the hypothesis that intrinsic or acquired defects in insulin biosynthesis characterize states of glucose intolerance, that these defects are worsened by excess lipid availability leading to a decreased size of the readily releasable pool of insulin. Incretin action may preserve beta cell function by maintenance of the synthetic rates sufficient to meet metabolic demands. Insulin biosynthetic rates will be measured for the first time in subjects with normal and impaired glucose tolerance by the novel method of mass isotopomer distribution analysis. Lay Summary: Controversy exists whether in those at risk for Type 2 Diabetes, there is an inability of pancreatic beta cells to make enough insulin or there is simply a failure to recognize glucose and appropriately release insulin. This proposal will measure for the first time the synthesis of insulin in normal subjects, those at risk, and those with Type 2 Diabetes. We will further determine whether the failure to handle increased fat exposure with more insulin insulin secretion in those at high risk of diabertes is due to decreased insulin synthesis, lack of adequate insulin stores, or an inability to release insulin in a timely fashion.